Due to the advancements in the field of optical metrology, it has found its applications in various areas such as biomedical, automotive, semiconductors, aerospace, etc. The popularity of optical techniques for metrology has increased by multiple folds owing to its non-invasive nature with ease of setup, fast data acquisition, and remote sensing ability. Optical techniques include hologram interferometry, speckle photography, speckle interferometry, Moire interferometry, photoelasticity, fringe projection technique, etc. The holographic interferometry technique works by quantifying the optical phase of the object by measuring the change in the interference fringes due to the shape of an object. This technique has a large number of advantages, but a steep object leads to a large number of fringes in the field of view, which are not resolvable as they fail to satisfy the Nyquist criterion. In this work, the fringe projection technique, which is a non-interferometric, non-invasive technique for generating 3D surface information is employed to measure the shape of a phase object like a wedge. Fringe projection is presented as a robust and compact technique for shape measurement of phase objects as it utilizes lesser components and has less complexity compared to the holographic technique.
Time multiplexing is a super resolution technique that sacrifices time to overcome the resolution reduction obtained because of diffraction. In structured illumination super-resolved imaging, high resolution and time changing patterns are projected on top of an object and a set of low resolution images are captured with the low quality imaging system. The set of low resolution images are digitally decoded with decoding patterns that are based on the high resolution projected encoding patterns.
In conventional structured-illuination approaches, the projected encoding patterns need to be a priori known in order to be used in the decoding process.
In this presentation, we will describe an enhancement of structured illumination approach towards label free imaging while obtaining the super resolved result without the need of requiring the a priori knowledge on the projected encoding patterns.
One of the drawbacks of using Lloyd’s mirror interferometer for quantitative phase contrast microscopy is that it leads to overlapping of the object information which restricts the use of field of view. One way to overcome this limitation is to prepare the sample slide in such a way that the object information is contained in only one portion of the illuminating beam, while the other acts as a separate reference beam. This geometry has excellent temporal stability and is used to examine human erythrocytes.
Digital holography interferometry (DHI) and digital holographic interference microscopy (DHIM) are tools that provide whole field information of the wavefront interacting with the object. This imaging modality can be an ideal tool for quantitative phase imaging of technical objects such as semiconductor samples. The phase information numerically reconstructed from the holograms can lead to extraction of the thickness/height of the sample. Usually DHI and DHIM are used in transmission mode for determination of optical thickness of the sample under investigation. In reflection mode this imaging techniques provide height of the sample.
For the semiconductor industry determination of height/thickness of object structures as well as the quantification of defects in the object structures is an important issue. The thickness and defect determination can be that of semiconductor thin films, micro/nano-pillars, LED displays, liquid crystal panels, the cover glasses used for protection of these structures etc. Digital holographic interferometric method (both reflection and transmission) can be used to design devices that can act as a fast, single shot technique for quantitative phase imaging of such samples. Such devices can provide more information about the sample compared to intensity based measurement systems. Also compact the digital holographic interferometric systems can be deployed in the manufacturing line of such devices to provide real time information.
We are involved in the design and development of digital holographic devices for inspection of semi-conductor wafers, thin films, displays and glass plates covering such samples. We have implemented digital holographic devices in the lens-less mode (in the case of DHI) and also with the use of an imaging lens (in the case of DHIM) both in reflection and transmission mode. DHI provides field of view equivalent to the sensor size, while DHIM technique was implemented with different magnifications, thereby providing varying field of views of the sample. Also in the case of DHI a propagation from the hologram plane (the plane at which the digital array for recording the hologram was situated) to the best focus plane (object plane) was realized by numerical implementation of diffraction integral. In DHIM, the digital array used for recording the hologram was at the image plane of the magnifying/de-magnifying lens. So the whole numerical reconstruction process reduced to Fourier fringe analysis, making the technique less computationally exhaustive, fast and quasi real-time. The developed devices were calibrated using known objects and then tested on different samples. The obtained results are found to be encouraging. In this paper, we describe our efforts in design, development and fabrication of digital holographic devices for the inspection of semiconductor samples.
Nonuniform refractive index distributions in transparent mediums are of interest as it gives rise to a modification of the probe light beam passing through such mediums. Various properties of the probe beam can be used to quantify the modification happening to the probe beam. One of these properties is the deflection of the beam. This could be used to map and quantify the spatiotemporal evolution of refractive index distribution in such mediums. The deflections could be measured by imaging the deflection of structured line pattern projected through such a system. We describe the development of a compact, portable device for mapping of refractive index distributions as well measurement of the diffusion coefficient of liquid solutions. The method and device are demonstrated by the real-time display of the refractive changes as well as measurement of diffusion coefficients in diffusing binary liquid solutions.
Quantitative three-dimensional (3-D) imaging of living cells provides important information about the cell morphology and its time variation. Off-axis, digital holographic interference microscopy is an ideal tool for 3-D imaging, parameter extraction, and classification of living cells. Two-beam digital holographic microscopes, which are usually employed, provide high-quality 3-D images of micro-objects, albeit with lower temporal stability. Common-path digital holographic geometries, in which the reference beam is derived from the object beam, provide higher temporal stability along with high-quality 3-D images. Self-referencing geometry is the simplest of the common-path techniques, in which a portion of the object beam itself acts as the reference, leading to compact setups using fewer optical elements. However, it has reduced field of view, and the reference may contain object information. Here, we describe the development of a common-path digital holographic microscope, employing a shearing plate and converting one of the beams into a separate reference by employing a pin-hole. The setup is as compact as self-referencing geometry, while providing field of view as wide as that of a two-beam microscope. The microscope is tested by imaging and quantifying the morphology and dynamics of human erythrocytes.
Adequate supply of oxygen to the body is the most essential requirement. In vertebrate species this function is performed by Hemoglobin contained in red blood cells. The mass concentration of the Hb determines the oxygen carrying capacity of the blood. Thus it becomes necessary to determine its concentration in the blood, which helps in monitoring the health of a person. If the amount of Hb crosses certain range, then it is considered critical. As the Hb constitutes upto 96% of red blood cells dry content, it would be interesting to examine various physical and mechanical parameters of RBCs which depends upon its concentration. Various diseases bring about significant variation in the amount of hemoglobin which may alter certain parameters of the RBC such as surface area, volume, membrane fluctuation etc. The study of the variations of these parameters may be helpful in determining Hb content which will reflect the state of health of a human body leading to disease diagnosis. Any increase or decrease in the amount of Hb will change the density and hence the optical thickness of the RBCs, which affects the cell membrane and thereby changing its mechanical and physical properties. Here we describe the use of lateral shearing digital holographic microscope for quantifying the cell parameters for studying the change in biophysical properties of cells due to variation in hemoglobin concentration.
Imaging and measurement of diffusion process in liquid solutions is a challenging and interesting problem. Especially the mixing of binary liquid solutions in real-time provides an insight into the physics of diffusion as well as leads to measurement of diffusion coefficient, which is the most important parameter of a diffusing liquid solution. Accurate measurement of diffusion coefficient is important in areas ranging from oil extraction to pollution control. Interferometric methods provides very accurate measurement of diffusion coefficients albeit they impose very stringent optical conditions. Here we describe the development of a compact, easy to implement, easy to use and inexpensive device for imaging and quantification of the diffusion process. This technique does not require the stringent optical conditions of interferometric techniques. It computes the diffusivity values by measuring the amount of deflection happening to a line pattern printed on a paper and projected through the sample cell. The measured diffusivity values varied by less than 1%, with the values of diffusivities reported in literature.
Measurement of rotation of plane of polarization of linearly polarized light can provide information about the concentration of the optically active system with which it interacts. For substances containing sugar, accurate measurement of rotation of linearly polarized light can provide quantitative information about concentration of sugar in the material. Measurement of sugar concentration is important in areas ranging from blood sugar level measurement in body fluids to measurement of sugar concentrations in juices and other beverages. But in many of these cases, the changes introduced to the state of polarization considering a sample of practical proportion is low and the measurement of low optical rotations becomes necessary. So methods with higher sensitivity, accuracy and resolution need to be developed for the measurement of low optical rotations. Here we describe the development of a compact, low cost, field portable, device for rotation sensing leading to sugar concentration measurements, using speckle de-correlation technique. The developed device measures rotations by determining the changes occurring to a speckle pattern generated by a laser beam passing through the medium under investigation. The device consists of a sample chamber, a diode laser module, a ground glass diffuser and a digital sensor for recording of laser speckle patterns. The device was found to have high resolution and sensitivity.
Digital holographic microscope is an ideal tool for quantitative phase contrast imaging of living cells. It yields the thickness distribution of the object under investigation from a single hologram. From a series of holograms the dynamics of the cell under investigation can be obtained. But two-beam digital holographic microscopes has low temporal stability due to uncorrelated phase changes occurring in the reference and object arms. One way to overcome is to use common path techniques, in which, the reference beam is derived from the object beam itself. Both the beams travel along the same path, increasing the temporal stability of the setup. In self-referencing techniques a portion of the object beam is converted into reference beam. It could be achieved by example, using a glass plate to create two laterally sheared versions of the object beam at the sensor, which interfere to produce the holograms/interferograms. This created a common path setup, leading to high temporal stability (~0.6nm). This technique could be used to map cell membrane fluctuations with high temporal stability. Here we provide an overview of our work on the development of temporally stable quantitative phase contrast techniques for dynamic imaging of micro-objects and biological specimen including red blood cells.
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